The Effect of Ovariectomy and Orchiectomy on Orthodontic Tooth Movement and Root Resorption in Wistar Rats.

STATEMENT OF THE PROBLEM
Root resorption (RR) after orthodontic tooth movement (OTM) is known as a multifactorial complication of orthodontic treatments. Hormonal deficiencies and their effect on bone turnover are reported to have influences on the rate of tooth movement and root resorption.


PURPOSE
This study was designed to evaluate the effect of female and male steroid sex hormones on tooth movement and root resorption.


MATERIALS AND METHOD
Orthodontic appliances were placed on the right maxillary first molars of 10 ovariectomized female and 10 orchiectomized male Wistar rats as experimental groups and 10 female and 10 male healthy Wistar rats as control groups. NiTi closed-coil springs (9mm, Medium, 011"×.030", Ortho Technology(®); Tampa, Florida) were placed between the right incisors and the first right maxillary molars to induce tipping movement in the first molars with the application of a 60g force. After 21 days, the rats were sacrificed and tooth movement was measured by using a digital caliper (Guanglu, China). Orthodontic induced root resorption (OIRR) was assessed by histomorphometric analysis after hematoxylin and eosin staining of sections of the mesial root.


RESULTS
The rate of tooth movement was significantly higher in all female rats, with the root resorption being lower in the experimental group. The rate of tooth movement in experimental male rats was significantly higher than the control group (p= 0.001) and the rate of root resorption was significantly lower in the experimental group (p= 0.001).


CONCLUSION
It seems that alterations in plasma levels of estrogen, progesterone, and testosterone hormones can influence the rate of OTM and RR. The acceleration in tooth movement increased OTM and decreased RR.


Introduction
Orthodontic tooth movement (OTM), as an essential component of orthodontic treatment, is achieved by bone remodeling during force application. Orthodonticinduced root resorption (OIRR) is an undesirable pathological consequence of orthodontic treatments, believed to be unavoidable, as in histological studies occurrence of greater than 90% of orthodontically induced inflammatory root resorption has been reported. [1][2][3][4] OIRR is usually asymptomatic and minimal with no clinical importance; however in rare cases, severe root resorption (more than ¼ of the root length) ends in root shortening and weakening of the tooth structure. [3,[5][6] During orthodontic force application, tooth movement occurs as a result of alveolar bone resorption due to the osteoclastic activity on the pressure side and bone deposition due to the osteoblastic induction on the tension side. [6][7] The periodontal ligament cells on the pressure side go through a necrosis process and after the hyaline zone formation, the tooth movement stops. The imbalance between osteoblastic and osteoclastic activities on the pressure and tension sides may result in resorptive areas on the root. [3,6,8] During the elimination process of the hyaline zone, mononuclear macrophages and multinuclear gigantic cells may damage the outer layer of the root which consists of cementoblasts. In more severe cases, the dentin may be affected as well. The resorption in dentin is irreversible and is considered a severe damage to the tooth structure. [5][6] The risk of OIRR is affected by many factors related to the patient and the orthodontic treatment.
From all patient-related factors, previous studies demonstrated that systemic factors, including hormone deficiencies and alveolar bone density, could influence OIRR. [1,[3][4]6] The rate of OTM is also affected by many factors including bone turnover. [9] Sex hormones like many other hormones have an impact on bone turnover. The influences of estrogen, progesterone, and testosterone on bone metabolism have been reported in many studies. [7,[10][11] The osteoporosis induced by estrogen and progesterone deficiencies in post-menopausal period results in bone loss and increased bone turnover. [12] Estrogens and androgens have been traditionally known as regulators of bone turnover in women and men. [8] The osteoporosis followed by the estrogen and progesterone withdrawal, affects the alveolar bone, promotes periodontal diseases and is a main cause of tooth loss. In orthodontic treatments, the increased bone turnover induced by lack of these hormones, results in progression of the tooth movement in an unstable pattern. [9,11,13] Osteoporosis and loss of bone mineral density caused by aging in male patients have also become an issue recently. [14][15] Osteoporosis in aged males due to lower androgen levels and the effect of androgens on bone density have been subjects of consideration in recent studies. [14,16] In males, androgens have been shown to be essential for skeletal maturation as well as maintaining the skeletal mass and bone density during mature stages. [17][18][19] Altogether, the effect of steroid sex hormones on bone turnover is inevitable.
There were studies designed to evaluate the effect of different bone turnover conditions induced by hormonal changes on orthodontic tooth movements and root resorption following orthodontic treatments.
These conditions included hyperparathyroidism, [20] hyper and hypothyroidism, [21][22] lactating, [23] and ovariectomization. [2,9,24] The influence of castration before puberty was also investigated in skeletal maturation. [25][26] Increased bone turnover in osteoporotic ovariectomized rats resulted in greater rates of OTM; however, few studies have demonstrated conflicting results on the OIRR rate in cases of estrogen and progesterone deficiencies. [2,27] Therefore, the present study was designed to evaluate the effect of estrogen and progesterone deficiencies on OTM and OIRR of maxillary first molars 21 days after force application with coil springs in female castrated mature rats compared with healthy mature female samples. Moreover, as far as the authors are aware, there were no previous studies on the effects of androgen deficiencies concerning OTM and OIRR. Therefore, these issues were also evaluated and compared between healthy and testosteronedeficient mature male rats followed by castration 21 days after force application.

Materials and Method
Twenty female and 20 male post-pubertal Wistar rats, aged 60 days, were used in this study. Ethical clearance of this study was obtained according to the guidelines of Animal Committee in Dental Research Center of Shahid Beheshti University of Medical Sciences.
Ten female rats were randomly placed in the control group and the other ten were selected for ovariectomy as the experimental group. In the male samples, 10 rats were randomly assigned to orchiectomy and the rest were placed in the control group. All the rats were housed for 7 days in the same place with similar temperature and light conditions and fed with the same diet for adaptation with the environmental conditions.

Castration procedure
The rats were weighed with a digital weighing scale and  Unitek). The appliance was set to induce a 60 g-force.
The force applied was measured by a force gauge (Dentaurum; Ispringen, Germany). The appliance was not reactivated during the experience and was designed to induce a tipping movement in the first maxillary molar.

OTM measurement
Twenty-one days after force application, the rats were sacrificed by an overdose of ketamine hydrochloride.
The distance between the distal surface of the first and mesial surface of the second right maxillary molar was measured by the same examiner with an electric caliper accurate to 0.001 mm (Tide Machine Tools Supply Co.; Shanghai, China).

OIRR and histomorphometric analysis
The maxillary jaws were excised and after 7 days of decalcification, the first molars were embedded in paraf-

Results
The hormone levels in the control groups at baseline and after 4 weeks were evaluated. The mean hormone levels in the experimental groups were measured before castration and 4 weeks after the operation. The descriptive mean values are presented in Tables 1 and 2.
The independent t-test showed significant differences in the mean progesterone and estrogen levels before and after ovariectomy (p= 0.001). The results of tooth movement and root resorption were compared with independent t-test and significant differences were detected between the control and experimental groups in male and female rats (p=-0.001). Tooth mo- The mean values of tooth movement and root resorption followed by orthodontic forces are summarized in Table 3.  Testosterone deficiency is a known risk factor for osteopenia and osteoporosis in older men. Less is known about the impact of testosterone deficiency on bone mineral density in younger men. The study of Kacker et al. reported that more than one third of men younger than 50 years with testosterone deficiency and infertility or sexual dysfunction had decreased bone mineral density and testosterone treatment increased the bone mineral density. [44] These